This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Disturbances in circadian rhythm expression appear to be associated with many diseases, such as seasonal affective disorders, a variety of other mental illnesses, sleep problems, and various metabolic syndromes. Even though associations between circadian rhythm disturbances and several diseases have been reported, the precise mechanisms by which circadian rhythm expression impacts the expression of diseases remain unknown. Among the limiting factors are technical difficulties related to the measurement of human circadian rhythm expression using physiological measures. To better understand human circadian rhythm expression in those patients and the implications of circadian rhythm disturbances in pathogenesis, as well as to evaluate potential pharmaceutical treatments, it is important to develop a non-invasive, simple, rapid and reliable in vitro assay to determine circadian rhythm expression in humans. One recently-developed technique allows us to obtain high-quality circadian rhythm records in real time, using the circadian clock-luciferase reporter gene and a lentivirus. Furthermore, researchers have demonstrated the feasibility of using skin fibroblasts to record human circadian rhythm expression. However, skin biopsy is not a routine procedure in most clinical departments, and they are painful for patients. Therefore, it would be beneficial to develop an assay system using some other cell types. Our preliminary data suggest that macrophages may be useful in this regard. In the present proposal, we aim to develop a non-invasive, in vitro assay system to measure human circadian rhythm in macrophages derived from human blood samples, using real-time monitoring of circadian parameters with a clock gene reporter (Aim 1). In addition, this technique will be applied to evaluate the effects of chemicals, thereby testing the feasibility of this assay (Aim 2) are confident to establish such an assay system using human blood cells, and determine the effects of various chemicals on circadian rhythm expression.